In Vitro Effects of Ketotifen and Cromolyn Sodium on Promastigotes and Amastigotes of Leishmania major
The first line treatment against cutaneous leishmaniasis is meglumine antimoniate. This drug is expensive and has serious side effects, including development of drug resistance.
In this research, because of paucity of information, the apoptotic and leishmanicidal effects of ketotifen and cromolyn sodium, as cell membrane stabilizer drugs, were investigated on standard strain of Leishmania major.
In this experimental study, L. major parasites were first cultured in RPM1 1640 media, supplemented with 10% fetal bovine serum (FBS) and antibiotics at 24 ± 1ºC. Drug concentrations of 5, 10, 15, and 20 µg/mL were then added to L. major culture at 24-, 48- and 72-hour intervals. The 3 - (4,5-dimethylthiazol-2-yl) -2,5-diphenyltetrazolium bromide (MTT) tetrazolium assays were performed to determine parasite viability and drug toxicity. Leishmania major promastigotes were augmented to the in vitro cultured macrophages (J774 cells) and then incubated for 72 hours. Half maximal inhibitory concentration (IC50) were ascertained by counting the parasites. The inhibitory effect of the drugs were compared with that of glucantime. Flow cytometry was performed in the next step, to evaluate apoptosis. Each test was repeated three times.
IC50 values of ketotifen and cromolyn sodium after 72 hours were calculated to be 2.04 and 17.67 µg/mL for promastigotes and 0.12 and 14.79 µg/mL for amastigotes, respectively. The results of MTT assays showed 20% and 35% promastigote viability after 72 hours of exposure to ketotifen and cromolyn sodium at 20 µg/mL concentration. Apoptosis in ketotifen and cromolyn sodium was quantified to be 11.52% and 9.96% in promastigotes and 99.5% and 98.6% in amastigote-infected macrophages, respectively. The results indicated that the drugs induce early and late apoptosis in parasites. All treatments produced results, which differed significantly from the control groups (P < 0.05).
Drugs used in this study, especially Ketotifen, showed lower toxicity yet similar anti-leishmanial effects on both forms, as cromolyn sodium did. It could be suggested that further investigations about the in vivo effects of these drugs, as candidates for cutaneous leishmaniasis treatment, are required.
Article Type:
Research/Original Article
Jundishapur Journal of Microbiology, Volume:12 Issue: 1, 2019
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